T Cell Memory to Pathogens: Generation and Function. We propose an integrated program to analyze CD4 and CD8 T cell memory against the respiratory pathogens causing influenza (flu) and tuberculosis (TB). In sophisticated animal models, we will define subsets of CD4 and CD8 memory T cells and determine the cellular and molecular basis of their generation, functions, location, persistence and relationship to other T cell subsets. This will greatly extend our basic understanding of immunity. "Generation and Persistence of CD4 Memory Subsets" (Swain), will determine the relationships of different functional subsets of CD4 T cells and will determine if they become functionally committed subsets of memory and define their protective functions. They will separate subsets of effector and memory CD4 T cells that work by killing infected targets and compare them to those that cause inflammation and secrete an inflammatory factor IL-17, and those that act by helping B cells. "CD8 Memory T Cells Mechanisms of Protection" (Dutton) will identify the mechanisms used by the CD8 T cell subsets that parallel those in Project 1 for CD4 subsets. In particular they will study the IL-17 producing subset of CD8 T cells that their preliminary data show plays a key role in protection against influenza, and compare these Tc17 to Tc1 subsets and define their function and protective abilities and mechanisms of action. "Regulation Of T Cell Homeostasis and Memory" (Bradley), will determine whether signals from "selectins" expressed on lung cells control CD4 and CD8 T cell responses and are needed for the development and persistence of CD4 memory cells. They will determine if selectin binding capacity identifies a distinct functional subset of CD4 cells, and what CD4 responses are selectin-dependent. "T Cell Memory to TB in the Lung" (Cooper), will determine factors that regulate induction of protective memory CD4 T cells, especially of the Th17 subset. They will also examine factors in the lung that regulate expression of memory T cell function in the lung and determine whether modulating the IL-17 memory response in the lung can increase protection. Defining the mechanisms by which memory T cell subsets provide protective immunity, is likely to result in identification of new correlates of protection for flu and TB, that will inform future vaccines targeted towards inducing robust T cell memory in addition to antibody, so that immunization will be effective even when new strains of TB and flu, including pandemic flu, emerge. Relevance: We believe that identifying the T cell subsets that contribute to protection and by defining mechanisms that regulate their generation and persistence, novel insights relevant to new strategies for improved vaccines will be obtained. PROJECT 1: Generation and persistence of CD4 memory subsets (Susan L. Swain, Project Leader) PROJECT 1 DESCRIPTION (provided by applicant): Generation and Persistence of CD4 Memory Subsets Recently we found that flu-specific CD4 T cell effectors provide protection against lethal challenge by virtue of both direct cytolytic activity (ThCTL) and help for Ab, suggesting additional functional heterogeneity among CD4 subsets that may contribute to CD4 memory heterogeneity and help us uncover new mechanisms of vaccine-induced protection. We have recent evidence that IL-17 producing CD4 also play a role in combating influenza. Our goal here is to evaluate the hypothesis that ThCTL cells represent a distinct functionally specialized subset that with Thi and Thi7 participate in an effective immune response. We will compare ThCTL to Thi7 and Thi subsets at the effector stage to further define their respective functions and determine how they provide protection in the lung against influenza. We will ask if ThCTL and Thi7 give rise to committed memory subsets and determine their roles in combating influenza. To accomplish these aims we will: 1) Isolate defined CD4 effector subsets generated in vitro and develop a scheme to separate them when they develop in vivo. We will define their cytokine and chemokine profiles, their migration and their involvement in help, cytotoxicity and protection against challenge with influenza. We will use the functional profile to develop a "signature" for each subset;2) We will determine each subset's ability to give rise to memory cells that respond to challenge and develop into secondary effectors and ask if they retain the same functional potential as they progressively differentiate;and 3) We will evaluate the mechanisms used by each memory Th subset to provide protection against lethal flu challenge. We will collaborate with: 1) Project 2 to determine which how the CD4 subsets compare with parallel CDS effector and memory subsets;2) Project 3 to determine migration of the CD4 subsets, and behavior in the lung and how interactions with selectins regulates function and memory generation from effectors;3) Project 4 to investigate whether similar Tuberculosis Ag-specific CD4 subsets can improve protecfion against Tuberculosis infection.